Fig. 6. GBM neural stem-like cells are the responding cells for endo-transdifferentiation by ETV2 expression in GBM patients.

a–c A fraction of GBM neural stem cells (nestin+/CD133+) co-expressed with CD31. a HE staining of a p-GBM paraffin section showing the GBM blood vessels (red box, a). Areas in dotted boxes are magnified below; CD31+ neural stem cells were detected by IF staining: CD133^high/nestin^high CD31^low Cells (1, arrows), CD133+/nestin+ CD31+ cells (2, arrows), CD133^low/nestin^low CD31^high Cells (3, arrows). b Coexpression of CD31 with mature neural maker genes (GFAP and NeuN) was rarely observed. Dashed circles highlight CD31+ cell clusters, arrows indicate NeuN+ CD31+ cells. c The nestin+ CD31+ ETV2+ triple-positive GBM cells were consistently detected. d ETV2 induced an endothelial signature in GBM neural stem-like cells. GBM neural stem-like cells (sox2+ nestin+, passage 3) were propagated as neurospheres under optimal conditions for neural stem cells. Flow cytometry, RT-PCR, and qPCR tests after 1–3 weeks of ETV2 overexpression indicated the successful induction of endothelial genes in GBM #109 neural stem-like cells. e Differentiated GBM tumor cells did not respond to ETV2 overexpression, and differentiated p-GBM tumor cells (GFAP+ NeuN+, passage 3) cultured with serum; RT-PCR and WB assays indicated that the differentiated GBM tumor cells (#109, #723, #24, at 2 weeks post LV-ETV2 infection) were unresponsive to ETV2 overexpression; HUVECs were set as the positive control. Scale bars in a, b, c, d, and e: 50 μm